|Title:||Functional studies of PfCRT related to chloroquine resistance in Plasmodium falciparum in Pichia pastoris|
|Subject:||Hong Kong Polytechnic University -- Dissertations|
|Department:||Department of Applied Biology and Chemical Technology|
|Pages:||xvii, 203 leaves : ill. (some col.) ; 30 cm|
|Abstract:||Malaria caused by protozoa of the genus Plasmodium is one of the most severe parasitic diseases worldwide. Malarial treatment has been hindered since the emergence of chloroquine resistance. The detailed resistance mechanism is still obscure and there are several proposed resistance mechanisms. PfCRT, a membrane protein found on the digestive vacuole of the parasite, has been demonstrated to cause chloroquine resistance despite its unknown functions. It is proposed that PfCRT has some pH regulatory functions which directly lead to chloroquine resistance. The pH of food vacuole, where chloroquine accumulates and acts, might be important in chloroquine resistance. According to the weak base theory, chloroquine accumulates less if the pH of food vacuole increases due to its weak base nature. PfCRT may cause chloroquine resistance by raising food vacuole pH. Besides, the sequence homology between PfCRT and the drug/metabolite transporter (DMT) superfamily implicates other transport functions of PfCRT. In this study, microsomes purified from P. pastoris expressed with PfCRT were employed as an in vitro system to characterize PfCRT. Its influences on microsomal pH were monitored by a pH sensitive fluorescence dye, BCECF, and the transport activity on a potential substrate, Hoechst 33342, was examined. Results showed that microsomes containing PfCRT (K76T or K76I) from resistant cell line have a significantly more acidic pH (ΔpH~ 0.13) which could be further acidified by addition of ATP or mefloquine. PfCRT could also initiate verapamil-reversible Hoechst 33342 transport in the presence of ATP or drugs. Chloride ions were found to be essential for PfCRT activity in both assays. These results indicate that PfCRT may be a pH-regulatory transporter which may alter pH through co-transporting its substrates with protons. The influences of ATP and chloride ions in both assays suggest there might be other proteins cooperating with PfCRT.|
|Rights:||All rights reserved|
Files in This Item:
|b17938016.pdf||For All Users||8.73 MB||Adobe PDF||View/Open|
As a bona fide Library user, I declare that:
- I will abide by the rules and legal ordinances governing copyright regarding the use of the Database.
- I will use the Database for the purpose of my research or private study only and not for circulation or further reproduction or any other purpose.
- I agree to indemnify and hold the University harmless from and against any loss, damage, cost, liability or expenses arising from copyright infringement or unauthorized usage.
By downloading any item(s) listed above, you acknowledge that you have read and understood the copyright undertaking as stated above, and agree to be bound by all of its terms.
Please use this identifier to cite or link to this item: